Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P04637 (p53)
 77,613 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previous work has shown that a fusion protein bearing a "nonremovable" N-terminal ubiquitin (Ub) moiety is short-lived in vivo, the fusion's Ub functioning as a degradation signal. The proteolytic system involved, termed the UFD pathway (Ub fusion degradation), was dissected in the yeast Saccharomyces cerevisiae by analyzing mutations that perturb the pathway. Two of the five genes thus identified, UFD1 and UFD5, function at post-ubiquitination steps in the UFD pathway. UFD3 plays a role in controlling the concentration of Ub in a cell: ufd3 mutants have greatly reduced levels of free Ub, and the degradation of Ub fusions in these mutants can be restored by overexpressing Ub. UFD2 and UFD4 appear to influence the formation and topology of a multi-Ub chain linked to the fusion's Ub moiety. UFD1, UFD2, and UFD4 encode previously undescribed proteins of 40, 110, and 170 kDa, respectively. The sequence of the last approximately 280 residues of Ufd4p is similar to that of E6AP, a human protein that binds to both the E6 protein of oncogenic papilloma viruses and the tumor suppressor protein p53, whose Ub-dependent degradation involves E6AP. UFD5 is identical to the previously identified SON1, isolated as an extragenic suppressor of sec63 alleles that impair the transport of proteins into the nucleus. UFD5 is essential for activity of both the UFD and N-end rule pathways (the latter system degrades proteins that bear certain N-terminal residues). We also show that a Lys --> Arg conversion at either position 29 or position 48 in the fusion's Ub moiety greatly reduces ubiquitination and degradation of Ub fusions to beta-galactosidase. By contrast, the ubiquitination and degradation of Ub fusions to dihydrofolate reductase are inhibited by the UbR29 but not by the UbR48 moiety. ufd4 mutants are unable to ubiquitinate the fusion's Ub moiety at Lys29, whereas ufd2 mutants are impaired in the ubiquitination at Lys48. These and related findings suggest that Ub-Ub isopeptide bonds in substrate-linked multi-Ub chains involve not only the previously identified Lys48 but also Lys29 of Ub, and that structurally different multi-Ub chains have distinct functions in Ub-dependent protein degradation.
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PMID:A proteolytic pathway that recognizes ubiquitin as a degradation signal. 761 50

Invasive cervical cancer is very highly correlated with the presence of high-risk human papillomavirus (HPV) types 16 and 18. Two viral proteins, E6 and E7, act in concert to subvert growth control of infected cells by inactivating the tumor suppressor proteins, p53 and Rb, respectively. E6 is thought to abrogate p53 function by stimulating its degradation via ubiquitin-mediated proteolysis in a reaction requiring E6AP (E6-Associated Protein). Here we evaluate the in vivo role of E6AP in p53 degradation in normal and HPV-infected cell types using antisense phosphorothioate oligodeoxynucleotides (S-ODNs). This study shows that reduction of E6AP in vivo in high-risk HPV-infected cells leads to an elevation of p53, confirming the function of E6AP predicted by in vitro experiments. Further, we demonstrate that reduction of E6AP in normal cells has no effect on p53 levels, indicative of an E6AP-indpendent mechanism for p53 degradation. These experiments show that inhibition of intermediate proteins in the ubiquitin-mediated proteolysis pathway (ubiquitin-conjugating enzymes or associated recognition proteins) can result in specific inhibition of substrate degradation. We propose that modulation of p53 levels by elimination of E6AP function may have therapeutic potential for cervical cancer.
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PMID:Antisense targeting of E6AP elevates p53 in HPV-infected cells but not in normal cells. 905 58

The cellular protein E6AP functions as an E3 ubiquitin protein ligase in the E6-dependent ubiquitination of p53. E6AP is a member of a family of functionally related E3 proteins that share a conserved carboxyl-terminal region called the Hect domain. Although several different E2 ubiquitin-conjugating enzymes have been shown to function with E6AP in the E6-dependent ubiquitination of p53 in vitro, the E2s that cooperate with E6AP in the ubiquitination of its normal substrates are presently unknown. Moreover, the basis of functional cooperativity between specific E2 and Hect E3 proteins has not yet been determined. Here we report the cloning of a new human E2, designated UbcH8, that was identified in a two-hybrid screen through specific interaction with E6AP. We demonstrate that UbcH7, an E2 closely related to UbcH8, can also bind to E6AP. The region of E6AP involved in complex formation with UbcH8 and UbcH7 was mapped to its Hect domain. Furthermore, we show that UbcH5 and UbcH6, two highly homologous E2s that were deficient for interaction with E6AP, could associate efficiently with another Hect-E3 protein, RSP5. Finally, only the E6AP-interacting E2s could function in conjunction with E6AP in the ubiquitination of an E6 independent substrate of E6AP, whereas the noninteracting E2s could not. Taken together, these studies demonstrate for the first time complex formation between specific human E2s and the Hect domain family of E3 proteins and suggest that selective physical interaction between E2 and E3 enzymes forms the basis of specificity for functionally distinct E2:E3 combinations.
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PMID:Physical interaction between specific E2 and Hect E3 enzymes determines functional cooperativity. 915 1

Common necessity for all papillomaviruses is to induce DNA synthesis in quiescent cells. This is commonly achieved by the E7 gene product, which interferes with the function of members of the retinoblastoma family controlling transition from the G1-phase to the S-phase of the cell cycle. Uncontrolled entry into S-phase activates, however, negative growth control signals which have to be bypassed to achieve production of progeny viruses. In addition to inherent activities of the E7 protein, high risk genital types encode an E6 protein that overcomes p53-mediated G1-arrest and apoptosis in concert with the cellular factor E6AP by targeting p53 for the enhanced ubiquitin-dependent degradation. The key question, which of these functions of genital E6 and E7 proteins is responsible for the carcinogenic phenotype, is still not completely answered. In contrast to high risk genital types no immortalizing or transforming activities have been found for the E7 proteins of the high risk cutaneous HPV8 and 47. On the other hand the ability of the E6 protein to transform established rodent fibroblasts seems to be a property shared by high risk genital and cutaneous types. To examine the existence of a common E6-mediated transforming pathway for both virus groups we compared the properties of the cutaneous E6 proteins with already known functions of E6 proteins of genital viruses. For this we analyzed the E6 proteins of low nak and high risk cutaneous and genital papillomaviruses with respect to cell transformation, to their abilities to bind, degradate, and influence the activity of human p53, and to bind E6AP. The results of our study demonstrate a clear lack of interaction between the transforming E6 proteins of HPV1 and HPV8 and both cellular proteins p53 and E6AP. In contrast, we found E6AP-independent binding of HPV16 E6 and HPV6 E6 to p53, although both proteins were different in their transforming potential. Of all four proteins investigated, only HPV16 E6 was able to bind to p53 and E6AP and to induce degradation of the p53 protein in the reticulocyte system. When we investigated in frame deletion mutants of the E6 protein of HPV16 for their abilities to bind to p53 or E6AP, degradate, and inhibit the transactivation function of p53 and to transform rodent fibroblasts, no correlation between the different activities could be found. Mutants still able to bind p53 and E6AP lacked transforming ability and other mutants that were transformation-competent were deficient in p53 and E6AP bindings.
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PMID:A comparative analysis of the interactions of the E6 proteins from cutaneous and genital papillomaviruses with p53 and E6AP in correlation to their transforming potential. 942 53

Human papillomavirus (HPV) E6 proteins inhibit apoptosis in both p53-dependent and p53-independent manners. A key point in apoptosis is the regulation provided by the Bcl-2 family; and in differentiating keratinocytes, in which HPV replicates, the Bak protein is highly expressed. We show that HPV-18 E6 will inhibit Bak-induced apoptosis and this is mediated by an interaction between the E6 and Bak proteins resulting in degradation of the Bak protein in vivo. We also show that Bak protein interacts with the ubiquitin ligase, E6AP, and that a mutant of Bak defective in E6AP binding is overexpressed in comparison with wild type. These studies suggest that Bak is probably the first naturally occurring target of E6AP to be identified.
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PMID:Inhibition of Bak-induced apoptosis by HPV-18 E6. 988 96

The E6AP ubiquitin-protein ligase (E3) mediates the human papillomavirus-induced degradation of the p53 tumor suppressor in cervical cancer and is mutated in Angelman syndrome, a neurological disorder. The crystal structure of the catalytic hect domain of E6AP reveals a bilobal structure with a broad catalytic cleft at the junction of the two lobes. The cleft consists of conserved residues whose mutation interferes with ubiquitin-thioester bond formation and is the site of Angelman syndrome mutations. The crystal structure of the E6AP hect domain bound to the UbcH7 ubiquitin-conjugating enzyme (E2) reveals the determinants of E2-E3 specificity and provides insights into the transfer of ubiquitin from the E2 to the E3.
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PMID:Structure of an E6AP-UbcH7 complex: insights into ubiquitination by the E2-E3 enzyme cascade. 1055 80

We previously showed that oncoprotein MDM2 has ubiquitin ligase activity toward tumor suppressor p53. In that paper, we showed very weak homology in the carboxyl terminal portion between MDM2 and E6AP (HECT domain). We mutated the cysteine residue (C464) corresponding to the residue essential for the ubiquitin ligase activity of E6AP and this mutation diminished the ligase activity of MDM2. The cysteine residue described above is also one of the cysteine residues that form the RING finger domain of MDM2. We tried to find out whether the diminishing of the activity by the mutation is attributable to the disruption of the RING finger domain or not. When the ring finger domain of MDM2 was deleted, the truncation mutant did not have the ubiquitin ligase activity. When we mutated the seven cysteine residues of RING finger domain of MDM2 in the carboxyl terminus, the disruption of each residue in the RING finger completely diminished the ubiquitin ligase activity of MDM2 toward MDM2 itself and toward tumor suppressor p53. These data indicate that the RING finger domain in MDM2 is essential for its ubiquitin ligase activity toward p53 and itself.
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PMID:Activity of MDM2, a ubiquitin ligase, toward p53 or itself is dependent on the RING finger domain of the ligase. 1072 39

The E6 protein of the high-risk human papillomaviruses (HPVs) and the cellular ubiquitin-protein ligase E6AP form a complex which causes the ubiquitination and degradation of p53. We show here that HPV16 E6 promotes the ubiquitination and degradation of E6AP itself. The half-life of E6AP is shorter in HPV-positive cervical cancer cells than in HPV-negative cervical cancer cells, and E6AP is stabilized in HPV-positive cancer cells when expression of the viral oncoproteins is repressed. Expression of HPV16 E6 in cells results in a threefold decrease in the half-life of transfected E6AP. E6-mediated degradation of E6AP requires (i) the binding of E6 to E6AP, (ii) the catalytic activity of E6AP, and (iii) activity of the 26S proteasome, suggesting that E6-E6AP interaction results in E6AP self-ubiquitination and degradation. In addition, both in vitro and in vivo experiments indicate that E6AP self-ubiquitination results primarily from an intramolecular transfer of ubiquitin from the active-site cysteine to one or more lysine residues; however, intermolecular transfer can also occur in the context of an E6-mediated E6AP multimer. Finally, we demonstrate that an E6 mutant that is able to immortalize human mammary epithelial cells but is unable to degrade p53 retains its ability to bind and degrade E6AP, raising the possibility that E6-mediated degradation of E6AP contributes to its ability to transform mammalian cells.
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PMID:Human papillomavirus type 16 E6 induces self-ubiquitination of the E6AP ubiquitin-protein ligase. 1086 52

The high-risk human papillomavirus (HPV) E6 proteins stimulate the ubiquitination and degradation of p53, dependent on the E6AP ubiquitin-protein ligase. Other proteins have also been shown to be targeted for degradation by E6, including hDlg, the human homolog of the Drosophila melanogaster Discs large (Dlg) tumor suppressor. We show here that the human homolog of the Drosophila Scribble (Vartul) (hScrib) tumor suppressor protein is also targeted for ubiquitination by the E6-E6AP complex in vitro and that expression of E6 induces degradation of hScrib in vivo. Characterization of the E6AP-E6-hScrib complex indicated that hScrib binds directly to E6 and that the binding is mediated by the PDZ domains of hScrib and a carboxyl-terminal epitope conserved among the high-risk HPV E6 proteins. Green fluorescent protein-hScrib was localized to the periphery of MDCK cells, where it colocalized with ZO-1, a component of tight junctions. E6 expression resulted in loss of integrity of tight junctions, as measured by ZO-1 localization, and this effect was dependent on the PDZ binding epitope of E6. Thus, the high-risk HPV E6 proteins induce the degradation of the human homologs of two Drosophila PDZ domain-containing tumor suppressor proteins, hDlg and hScrib, both of which are associated with cell junction complexes. The fact that Scrib/Vart and Dlg appear to cooperate in a pathway that controls Drosophila epithelial cell growth suggests that the combined targeting of hScrib and hDlg is an important component of the biologic activity of high-risk HPV E6 proteins.
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PMID:Human scribble (Vartul) is targeted for ubiquitin-mediated degradation by the high-risk papillomavirus E6 proteins and the E6AP ubiquitin-protein ligase. 1102 93

Recent analyses have identified a number of binding partners for E6, including E6AP, ERC55, paxillin, hDlg, p300, interferon regulatory factor 3, hMCM7, Bak, and E6TP1. Notably, association with E6 targets p53, E6TP1, myc, hMCM7, and Bak for degradation. However, the relative importance of the various E6 targets in cellular transformation remains unclear. E6 alone can dominantly immortalize normal human mammary epithelial cells (MECs), permitting an assessment of the importance of various E6 targets in cellular transformation. Studies in this system indicate that E6-induced degradation of p53 and E6 binding to ERC55 or hDlg do not correlate with efficient immortalization. Here, we have examined the role of E6TP1, a Rap GTPase-activating protein, in E6-induced immortalization of MECs. We tested a large set of human papillomavirus type 16 E6 mutants for their ability to bind and target E6TP1 for degradation in vitro and in vivo. We observed a strict correlation between the ability of E6 protein to target E6TP1 for degradation and its ability to immortalize MECs. Recent studies have identified telomerase as a target of E6 protein. Previous analyses of E6 mutants have revealed this trait to closely correlate with MEC immortalization. We examined our entire panel of E6 mutants for rapid induction of telomerase activity and found in general a strong correlation with immortalizing ability. The tight correlation between E6TP1 degradation and MEC immortalization strongly supports a critical role of functional inactivation of E6TP1 in E6-induced cellular immortalization.
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PMID:Human papillomavirus type 16 E6-induced degradation of E6TP1 correlates with its ability to immortalize human mammary epithelial cells. 1128 1


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